单胺氧化酶 A 抑制剂氯吉宁是广谱真菌 ABC 和 MFS 转运蛋白外排泵抑制剂，可逆转白色念珠菌和光滑念珠菌临床分离株的唑类耐药性。
The monoamine oxidase A inhibitor clorgyline is a broad-spectrum inhibitor of fungal ABC and MFS transporter efflux pump activities which reverses the azole resistance of Candida albicans and Candida glabrata clinical isolates.
机构信息
Sir John Walsh Research Institute and Department of Oral Sciences, University of Otago, Dunedin, New Zealand.
出版信息
Antimicrob Agents Chemother. 2012 Mar;56(3):1508-15. doi: 10.1128/AAC.05706-11. Epub 2011 Dec 27.
Resistance to the commonly used azole antifungal fluconazole (FLC) can develop due to overexpression of ATP-binding cassette (ABC) and major facilitator superfamily (MFS) plasma membrane transporters. An approach to overcoming this resistance is to identify inhibitors of these efflux pumps. We have developed a pump assay suitable for high-throughput screening (HTS) that uses recombinant Saccharomyces cerevisiae strains hyperexpressing individual transporters from the opportunistic fungal pathogen Candida albicans. The recombinant strains possess greater resistance to azoles and other pump substrates than the parental host strain. A flow cytometry-based HTS, which measured increased intracellular retention of the fluorescent pump substrate rhodamine 6G (R6G) within yeast cells, was used to screen the Prestwick Chemical Library (PCL) of 1,200 marketed drugs. Nine compounds were identified as hits, and the monoamine oxidase A inhibitor (MAOI) clorgyline was identified as an inhibitor of two C. albicans ABC efflux pumps, CaCdr1p and CaCdr2p. Secondary in vitro assays confirmed inhibition of pump-mediated efflux by clorgyline. Clorgyline also reversed the FLC resistance of S. cerevisiae strains expressing other individual fungal ABC transporters (Candida glabrata Cdr1p or Candida krusei Abc1p) or the C. albicans MFS transporter Mdr1p. Recombinant strains were also chemosensitized by clorgyline to other azoles (itraconazole and miconazole). Importantly, clorgyline showed synergy with FLC against FLC-resistant C. albicans clinical isolates and a C. glabrata strain and inhibited R6G efflux from a FLC-resistant C. albicans clinical isolate. Clorgyline is a novel broad-spectrum inhibitor of two classes of fungal efflux pumps that acts synergistically with azoles against azole-resistant C. albicans and C. glabrata strains.
由于 ATP 结合盒（ABC）和主要易化因子超家族（MFS）质膜转运蛋白的过度表达，对常用唑类抗真菌药物氟康唑（FLC）的耐药性可能会产生。克服这种耐药性的一种方法是鉴定这些外排泵的抑制剂。我们开发了一种适用于使用重组酿酒酵母菌株的高通量筛选（HTS）泵测定法，该菌株过度表达来自机会性真菌病原体白色念珠菌的单个转运蛋白。重组菌株对唑类药物和其他泵底物的抗性高于亲本宿主菌株。一种基于流式细胞术的 HTS 用于测量荧光泵底物罗丹明 6G（R6G）在酵母细胞内的细胞内保留增加，用于筛选 1200 种市售药物的 Prestwick 化学文库（PCL）。鉴定出九种化合物为命中化合物，单胺氧化酶 A 抑制剂（MAOI）氯丙嗪被鉴定为两种白色念珠菌 ABC 外排泵 CaCdr1p 和 CaCdr2p 的抑制剂。二次体外测定证实氯丙嗪抑制泵介导的外排。氯丙嗪还逆转了表达其他单个真菌 ABC 转运蛋白（光滑念珠菌 Cdr1p 或克鲁斯氏念珠菌 Abc1p）或白色念珠菌 MFS 转运蛋白 Mdr1p 的酿酒酵母菌株的 FLC 耐药性。重组菌株也被氯丙嗪对其他唑类药物（伊曲康唑和咪康唑）变得化学敏感。重要的是，氯丙嗪与 FLC 对 FLC 耐药的白色念珠菌临床分离株和光滑念珠菌菌株显示出协同作用，并抑制 FLC 耐药的白色念珠菌临床分离株中 R6G 的外排。氯丙嗪是两种真菌外排泵的新型广谱抑制剂，与唑类药物联合使用可对抗唑类耐药的白色念珠菌和光滑念珠菌菌株。
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